The present invention relates generally to methods for genetically altering higher plant materials and reproducing whole plants therefrom. More particularly, the invention relates to a method for genetically transforming pea plants.
The garden pea (Pisum sativum L.) is a legume and an important food crop. As with other legumes, pea plants are able to obtain nitrogen from symbiotic soil bacteria and have a substantially reduced fertilizer requirement compared to other non-leguminous crops. Thus, the production of new, improved cultivars of pea could be of great commercial value, particularly in view of increasing fertilizer costs.
As with any valuable plant species, breeders have long used conventional cross-breeding techniques to improve existing varieties and create new cultivars. While improvements have been achieved, cross-breeding techniques are laborious and slow because of the time required to breed and grow successive plant generations.
Conventional breeding methods can only utilize those genes that are present in species that are sexually compatible with P. sativum. Thus, it would be desirable to utilize recombinant DNA technology to produce new pea varieties and cultivars in a controlled and predictable manner that contain genes both from sexually compatible crops, and from other unrelated plants, animals, bacteria and viruses.
The recombinant DNA manipulation of pea, however, has been hindered by difficulty in regenerating whole plants from tissue culture, by difficulty in obtaining transformed pea tissue and ultimately, in linking regeneration with transformation.
For these reasons, it would be desirable to provide improved methods for the recombinant DNA transformation of pea plant material and the regeneration of whole plants from the transformed material. It would be particularly desirable to be able to introduce desired characteristic(s) to such material(s) and to be able to regenerate viable pea plantlets from the modified materials. Such methods should be capable of introducing a preselected exogenous gene(s) into the pea plant material and should permit the selection of transformed shoots which are regenerated from the material. The method should produce regenerated pea plants which have stably incorporated the gene(s).
2. Description of the Background Art
DeKathen et al. (1990) Plant Cell Rep. 9:276-279, describes the Agrobacterium-mediated transformation of epicotyl and nodus explants from Madria (an inedible pod variety of pea). Related work was described in DeKathen et al. (1990) J. Cell Biochem., Suppl. O (14 Part E), Abstract R125, p. 282. Puonti-Kaerlas et al. (1990) Theor. and Appl. Genet. 80:246-252, describes the regeneration of transformed shoots from transformed callus material of two inedible pod varieties of pea.
Jackson and Hobbs (1990) In Vitro Cell. Dev. Biol. 26:835-838, describes the regeneration of whole plants from pea plumule material, without transformation. PBI Bulletin, May 1991, NRC Plant Biotechnology Institute, National Research Council Canada, pp 2-4, asserts that the culture method of Jackson and Hobbs has been used for the transformation of pea with Agrobacterium vectors. See, Jordan et al. (1990) J. Cell Biochem., Suppl. O (14 Part E), Abstract R126, p. 282.
Transformation of pea plant material without whole plant regeneration is described in Lulsdorf et al. (1991) Plant Cell Rep. 9:479-483; Puonti-Kaerlas (1989) Plant Cell Rep. 8:321-324; Hussey et al. (1989) 148:101-105; Filipone and Lurquin (1988) Genet. Agrar. 42:452 (Abstract); and Nauerby et al. (1991) Plant Cell Rep. 9:676-679.
The transformation of legumes other than pea is described in Hinchee et al. (1988) Bio/Technology 6:915-922 (soybean); and Eapen et al. (1987) Theor. Appl. Genet. 75:207-210.